Biosensors directed towards disease and cancer detection often target biomarkers such as proteins that are not present in healthy individuals. The buildup of these biomarkers correlates with the progression of the particular disease. Unfortunately, the number of available treatment options and the overall prognosis decrease considerably as the disease progresses from early to advanced stages. Further, many diseases are asymptomatic until the advanced stages, at which point options are limited. For these reasons, early detection is critical for successful treatment.
The gold standard for biosensing has been ELISA (enzyme-linked immunosorbent assay), which is utilized in commercial products such as pregnancy tests, as well as for detection of antibiotics in milk and the presence of salmonella. Similarly, sandwich assays can be performed with fluorescently-labeled secondary antibodies. In both cases, however, antibody labeling is a requirement. While these labeled methods typically provide adequate signal-to-noise ratio, the production of labeled antibodies is expensive, time consuming, and can influence molecular binding. Further, since the sandwich assay requires multiple incubation steps, these assays do not permit real-time detection.
Many techniques have been developed to overcome the shortcomings of bio sensors that require molecular labeling. These techniques typically probe the interfacial binding of a particular protein by means of changes in interfacial refractive index or mass. Surface plasmon resonance (Knoll, W., Interfaces and thin films as seen by bound electromagnetic waves, Annual review of physical chemistry 1998, 49 (1), 569-638) and ellipsometry (Ostroff, R. M.; Maul, D.; Bogart, G. R.; Yang, S.; Christian, J.; Hopkins, D.; Clark, D.; Trotter, B.; Moddel, G., Fixed polarizer ellipsometry for simple and sensitive detection of thin films generated by specific molecular interactions: applications in immunoassays and DNA sequence detection, Clin Chem 1998, 44 (9), 2031-2035) are common techniques for monitoring interfacial refractive index, while quartz crystal microbalance (Liss, M.; Petersen, B.; Wolf, H.; Prohaska, E., An Aptamer-Based Quartz Crystal Protein Biosensor, Analytical chemistry 2002, 74 (17), 4488-4495) measures interfacial mass change. A number of different modifications have been proposed to reduce the noise in surface plasmon resonance (SPR) sensors such as polarization interferometry (Sun, Z. L.; He, Y. H.; Guo, J. H., Surface plasmon resonance sensor based on polarization interferometry and angle modulation, Applied Optics 2006, 45 (13), 3071-3076) and differential phase change (Wu, S.; Ho, H.; Law, W.; Lin, C.; Kong, S., Highly sensitive differential phase-sensitive surface plasmon resonance biosensor based on the Mach-Zehnder configuration, Optics letters 2004, 29 (20), 2378-2380). A very low (or possibly the lowest known) detection limit for SPR based biosensing has been demonstrated by Li et al. (Li, Y. C.; Chang, Y. F.; Su, L. C.; Chou, C., Differential-phase surface plasmon resonance biosensor, Analytical chemistry 2008, 80 (14), 5590-5595). Li et al. use differential-phase-sensitive surface plasmon resonance to monitor the interaction between mouse IgG and antimouse IgG at 67 attomolar concentration. While this is an impressive achievement, there are a number of disadvantages with the experimental setup of Li et al. One important disadvantage relates to the sensitivity of the apparatus. It is well known that for phase-sensitive SPR measurements, the largest sensitivity to phase change occurs at the minimum angle of reflectivity. However, operating at minimum reflection where only a small percentage of photons reach the detector severely reduces the signal-to-noise ratio. Operating outside of the minimum reflection or changing the gold thickness can increase reflectivity but only at the expense of the phase sensitivity. Increasing laser power is also not a viable option, since the adsorbed photons dissipate as heat in the dielectric material of interest and at high powers can create temperature gradients in the sample. Further, phase measurements are sensitive to the roughness of the gold film. Achieving ultrasmooth gold films and substrates for gold deposition can be a daunting task that is required for this configuration to maintain the optimum sensitivity. In addition, there is an extremely narrow dynamic range for phase detection. Thus, only extremely low interfacial concentrations or smaller molecules can be measured. For real-world applications, such as detection in serum, non-specific adsorption alone would likely cause departure from the usable detection range.